Proton ATPases are a large class of membrane-proteins that use the energy of ATP hydrolysis to generate an electrochemical proton gradient across a membrane. The resultant gradient may be used to transport other ions across the membrane (Na.sup.+, K.sup.+, or Cl.sup.-) or to maintain an acidic environment important to the function of many cellular vesicles (Mellman, I. et al. (1986) Ann. Rev. Biochem. 55:663-700). Proton ATPases are further subdivided into the mitochondrial F-ATPases, the plasma membrane ATPases, and the vacuolar ATPases.
The vacuolar proton ATPases (vp-ATPases) provide most of the energy required for transport processes in the vacuolar system in eukaryotic cells. They establish and maintain an acidic pH within various vesicles involved in the processes of endocytosis and exocytosis. Such vesicles include phagosomes, lysosomes, endosomes, and secretory vesicles. Endocytosis is the process in cells of internalizing nutrients, solutes or small particles (pinocytosis), or large particles such as internalized receptors, viruses, bacteria, or bacterial toxins (phagocytosis). Exocytosis is the process of transporting molecules to the cell surface. Exocytosis facilitates the placement or localization of membrane-bound receptors or other membrane proteins and the secretion of hormones, neurotransmitters, digestive enzymes, and wastes. Endocytosis and exocytosis are fundamental to the function of all types of cells.
The vp-ATPases and the F-ATPases, which function in ATP synthesis and hydrolysis in mitochondria, are related in both their subunit structure and evolutionary origin. Both contain distinct catalytic and membrane sectors, each sector containing multiple subunits. The catalytic sector of vp-ATPase consists of five subunits designated A (72 kDa), B (57 kDa), C (41 kDa), D (34 kDa), and E (33 kDa) (Nelson, H. et al. (1995) Proc. Natl. Acad. Sci. 92:497-501). Three subunits, ACI 15, AC39, and a proteolipid component, have been identified in the membrane sector of vp-ATPase from various sources. A fourth subunit, AC45, has also been found in vp-ATPase from bovine adrenal glands (Supek, F. et al. (1994) J. Biol. Chem. 269: 24102-6). The membrane sector has several functions, including proton conduction across the membrane, energy coupling with the catalytic sector, communication with the lumen, and modulation of enzyme activity. Mutational studies in yeast have shown that, while the membrane sector may be assembled independently of the catalytic sector, assembly of the catalytic sector is absolutely dependent on previous assembly of the membrane sector (Supek et al. supra; Noumi, T. et al. (1991) Proc. Natl. Acad. Sci. 88: 193842). Thus, expression and assembly of the membrane sector subunits control the overall activity of the enzyme complex.
The AC45 membrane subunit is a 468 amino acid protein with a calculated molecular mass of 51 kDa. A potential signal sequence comprises the first 35 amino acids, and a potential transmembrane domain has been identified at the C-terminus. The protein contains eight potential glycosylation sites between the N-terminal signal sequence and the transmembrane domain. Utilization of these glycosylation sites is indicated by the fact that the inclusion of dog pancreas microsomes during cell-free synthesis of the protein significantly decreased its electrophoretic mobility on SDS-polyacrylamide gel (Supek et al., supra).
The discovery of a new vacuolar ATPase AC45 subunit and the polynucleotides encoding it satisfies a need in the art by providing new compositions which are useful in the diagnosis, prevention, and treatment of cancer and immune disorders.